Protonation states of the catalytic intermediates of cytochrome c oxidase.

Protonation changes accompanying conversion of oxidised (O state) cytochrome c oxidase to the 2-electron-reduced P state, and 3-electron-reduced F state at pH 8.0 have been measured. It was found that 2 and 3 protons, respectively, were taken up. The fourth proton required for the reduction of O2 to H2O must therefore be consumed in the remaining F----O portion of the catalytic cycle.     

Sulfur dynamics in mineral horizons of two northern hardwood soils. A column study with 35S

Sulfur dynamics of two Spodosols were ascertained using soil columns constructed from homogenized mineral soil from nothern hardwood ecosystems at the Huntington Forest (HF) in the Adirondack Mountains of New York and Bear Brook Watershed in Maine (BBWM). Columns were leached for 20 weeks with a simulated throughfall solution with³⁵SO₄ ^2-. Sulfur constituents were similar to those of other Spodosols, with the organic S fractions (C-bonded S and ester sulfate) constituting over 90% of total S. HF soil columns had higher total S (14.9 mol S g^-1) than that for the BBWM soil columns (7.4 mol g^-1) primarily due to higher C-bonded S in the former.Initially, adsorbed SO₄ ^- accounted for 5 and 4% of total S for the BBWM and HF soil columns, respectively. After 20 weeks, adsorbed SO₄ ^2- decreased (81%) in BBWM and increased (33%) in HF soil columns. For both HF and BBWM soil columns, C-bonded S increased and ester sulfate decreased, but only for HF columns was there a net mineralization of organic S (5.6% of total S). The greatest decrease in ester sulfate occurred at the top of the columns.Leaching of³⁵S was less than 0.5% of the³⁵S added due to its retention in various S constituents. There was an exponential decrease in³⁵S with column depth and most of the radioisotope was found in C-bonded S (70–88 and 70–91% for BBWM and HF, respectively). The rapid turnover of adsorbed SO^2- ₄ was reflected in its high specific activity (834 and 26 kBq mol^-1 S for BBWM and HF, respectively). The lower specific activity of adsorbed SO₄ ^2- in HF was attributable to greater isotopic dilution by non-radioactive SO^2- ₄ derived from greater organic S mineralization in the HF versus the BBWM columns.Both soil columns initially had high levels of NO^- ₃ which resulted in the generation of H⁺ and net retention of SO₄ ^2- in the early phase of the experiment due to pH dependent sulfate adsorption; later NO₃ ^- decreased and SO₄ ^2- was desorbed. Leaching of NIO₃ ^- and SO₄ ^2- was correlated with losses of Mg²⁺ and Ca²⁺ of which the latter was the dominant cation.Analyses using both S mass balances and radioisotopes corroborate that for BBWM soil columns, SO^2- ₄ adsorption-desorption dominated the S biogeochemistry while in HF soil columns, organic S mineralization-immobilization processes were more important. It is suggested that similar techniques can be applied to soils in the field to ascertain the relative importances of SO₄ ^2- adsorption processes and organic S dynamics.     

Contraction of smooth muscle of pig airway tissues from before birth to maturity

Two heavy chains of smooth muscle myosin (MHC1 and MHC2) were identified in pig airways and parenchyma. The ratio of MHC1 to MHC2 was the same along the bronchial tree in animals of the same age, but it changed with age (mature, young, suckling, and fetus), ranging from 0.8 in the mature to 2.2 in the fetus. Stress developed in airway (trachea, bronchus, and bronchiole) and parenchymal preparations in response to carbachol and histamine (mN/mm2) was normalized for myosin content (N/mm2 myosin). Airways from sucklings always developed the greatest stress to carbachol and histamine with the rank order of maximum force (Emax) suckling greater than fetus greater than young greater than mature for carbachol in large airways. Airway ranking to histamine was similar except that Emax of fetal bronchus and bronchiole were least. In parenchymal strips, mature animals gave strong responses to carbachol and histamine compared with other age groups. Sensitivity to carbachol was increased in the suckling trachea; otherwise it did not vary with age. Chemically skinned tracheal fibers exhibited three- to fourfold greater sensitivity to Ca2+ in fetal and suckling airways compared with the older animals. It is concluded that maturation of smooth muscle occurs in the expression of myosin, in the Ca2(+)-force relationships of the contractile machinery, and in the pharmacological responsiveness of the intact smooth muscle, with the latter greatest at or soon after birth.     

Twenty-year outcome analysis of genetic screening programs for Tay-Sachs and beta-thalassemia disease carriers in high schools.

Programs for education, screening, and counseling of senior-high-school students, in populations at high risk for Tay-Sachs and beta-thalassemia diseases, have existed for >20 years in Montreal. Four process and outcome variables are reported here: (i) voluntary participation rates in the high-school cohort; (ii) uptake rates for the screening test; (iii) origin of carrier couples seeking the prenatal diagnosis option in the programs; and (iv) change in incidence of the two diseases. Between 1972 and 1992, we screened 14,844 Ashkenazi-Jewish students, identified 521 HexA-deficient carriers (frequency 1:28), reached 89% of the demographic cohort in the educational component of the program, and achieved 67% voluntary participation in the subsequent screening phase. The corresponding data for the beta-thalassemia program are 25,274 students (mainly of Mediterranean origin) representing 67% of the cohort with 61% voluntary participation in the screening phase (693 carriers; frequency 1:36). From demographic data, we deduce that virtually all the carriers identified in the high-school screening program remembered their status, had their partner tested if they did not already know they were a carrier couple, and took up the options for reproductive counseling/prenatal diagnosis. In Montreal, the current origin of all couples using prenatal diagnosis for Tay-Sachs and beta-thalassemia diseases is the corresponding genetic screening/testing program, whereas, at the beginning of the programs, it was always because there was a history of an affected person in the family. Incidence of the two diseases has fallen by 90%-95% over 20 years; the rare new cases are born (with two exceptions) outside the target communities or to nonscreened couples.     

Clustering of Marine Bacteria in Seawater Enrichments

Seawater enrichments of marine bacteria clustered in 20- to 50-(mu)m-wide bands near air-water interfaces. The cells within the band travelled at up to 212 (mu)m s(sup-1) and at an average speed of 163 (mu)m s(sup-1). Mean cell speeds peaked mid-run at 187 (mu)m s(sup-1). At the end of the run, bacteria reversed direction rather than randomly reorienting. The duration of the stops during reversal was estimated at 18 ms, six to seven times shorter than that found in enteric bacteria. Cells hundreds of micrometers from the band travelled at half the speed of the bacteria in the band. The fastest isolate from the seawater enrichment was identified as Shewanella putrefaciens and had an average speed of 100 (mu)m s(sup-1) in culture. Air-water interfaces produced no clustering or speed changes in isolates derived from enrichments. Salinity and pH, however, both influenced speed. The speed and reversal times of the seawater enrichments indicate that the bacteria in them are better adapted for clustering around small point sources of nutrients than are either enteric or cultured marine bacteria.     

A Second Gene for Cerulean Cataracts Maps to the β Crystallin Region on Chromosome 22

Congenital cataracts are one of the most common major eye abnormalities and often lead to blindness in infants. At least a third of all cases are familial. Within this group, highly penetrant, autosomal dominant forms of congenital cataracts (ADCC) are most common. ADCC is a genetically heterogeneous group of disorders, in which at least eight different loci have been identified for nine clinically distinct forms. Among these, Armitageet al.(Nature Genet.9: 37–40, 1995) mapped a gene for cerulean blue cataracts to chromosome 17q24. Bodkeret al.(Am. J. Med. Genet.37: 54–59, 1990) described a large family with cerulean blue cataracts, in which the affected daughter of affected first cousins was presumed to be homozygous for the purported gene. We report linkage in this family to the region on chromosome 22q that includes two β crystallin genes (CRYBB2, CRYBB3) and one pseudogene (CRYBB2P1). The affected female in question is homozygous at all markers.     

Analysis of RIM11, a yeast protein kinase that phosphorylates the meiotic activator IME1.

Many yeast genes that are essential for meiosis are expressed only in meiotic cells. Known regulators of early meiotic genes include IME1, which is required for their expression, and SIN3 and UME6, which prevent their expression in nonmeiotic cells. We report here the molecular characterization of the RIM11 gene, which we find is required for expression of several early meiotic genes. A close functional relationship between RIM11 and IME1 is supported by two observations. First, sin3 and ume6 mutations are epistatic to rim11 mutations; prior studies have demonstrated their epistasis to ime1 mutations. Second, overexpression of RIM11 can suppress an ime1 missense mutation (ime1-L321F) but not an ime1 deletion. Sequence analysis indicates that RIM11 specifies a protein kinase related to rat glycogen synthase kinase 3 and the Drosophila shaggy/zw3 gene product. Three partially defective rim11 mutations alter residues involved in ATP binding or catalysis, and a completely defective rim11 mutation alters a tyrosine residue that corresponds to the site of an essential phosphorylation for glycogen synthase kinase 3. Immune complexes containing a hemagglutinin (HA) epitope-tagged RIM11 derivative, HA-RIM11, phosphorylate two proteins, p58 and p60, whose biological function is undetermined. In addition, HA-RIM11 immune complexes phosphorylate a functional IME1 derivative but not the corresponding ime1-L321F derivative. We propose that RIM11 stimulates meiotic gene expression through phosphorylation of IME1.     

ASSORTATIVE MATING AND NATURAL SELECTION IN AN IRIS HYBRID ZONE

The phenology of different genotypes and the distribution of genetic variation among flowering plants and their progeny were examined to assess the levels of assortative mating and selection in a hybrid population of Iris. This study and a previous survey of RAPD nuclear markers and chloroplast markers indicate that the population consists of parental genotypes and recombinant hybrid genotypes that are similar to the parental species (I. fulva and I. brevicaulis), although lacking intermediate genotypes. Early in the season only I. fulva genotypes produced flowers, but as flowering in these plants decreased, the hybrid genotypes and I. brevicaulis genotypes began flowering, resulting in a 24-d period of coincidental flowering. The genotypic distribution of seeds produced during the period of flowering overlap contained a high frequency of intermediate genotypes that were not present in the adult generation. The degree of effective assortative mating was examined by comparing the observed progeny genotypic distributions with expected distributions from a mixed-mating model. The model included selfing and random outcrossing to the nearest plants that had pollen-bearing flowers on the day the recipient flower was receptive. The observed genotypic distribution of progeny from plants with I. brevicaulis chloroplast DNA (cpDNA) was not significantly different from the expected distribution. For I. fulva genotypes, however, there were higher than expected frequencies in the extreme genotypic classes, although intermediate genotypes were absent, indicating that these plants were preferentially mating with similar genotypes. Compared with the extreme genotypes, a larger proportion of the intermediate seed progeny produced were aborted, indicating that intermediate genotypes have lower viability. On the basis of the observed progeny genotypes and genetic disequilibria estimates for the adults and the progeny, there appears to be a pattern of effective asymmetrical mating in this population. This asymmetry is most likely due to pollen-style interactions that reduce the fertilization ability of genetically dissimilar pollen, or preferential abortion of genetically intermediate zygotes by I. fulva-like genotypes. The lack of any apparent discrimination by I. brevicaulis-like genotypes creates a directional exchange of nuclear genetic elements that will have implications for introgression and the evolution of hybrid genotypes.     

Mouse centromere mapping using oligonucleotide probes that detect variants of the minor satellite

Cytologically, the centromere is found at the very end of most Mus musculus chromosomes, co-localizing with an array of minor satellite sequences. It is separated from the euchromatin of the long arm by a large domain of heterochromatin, composed in part of arrays of major satellite sequences. We used oligonucleotide probes that specifically detect regions of sequence variation found in certain cloned minor satellite sequences. They detect a limited subset of the minor satellite arrays in the mouse genome, based on both pulsed-field gel electrophoresis and in situ hybridization data, and provide direct molecular genetic markers for individual centromeres in some inbred mouse strains. Array size polymorphisms detected by these probes map to positions consisten with the centromeres of chromosomes 1 and 14 in the BXD recombinant inbred (RI) strains. The genetic distances between these minor satellite arrays and loci on the long arms of chromosomes 1 and 14 are consistent with repression of meiotic recombination in the heterochromatic domains separating them. The existence of chromosome-specific minor satellite sequences implies that the rate of sequence exchange between non-homologous chromosomes relative to the rate between homologous chromosomes is much lower than has previously been postulated. We suggest that the high degree of sequence homogeneity of mouse satellite sequences may instead reflect recent common ancestry.     

Smilax saülensis (Smilacaceae), a new species from French Guiana

Botanical exploration of the vicinity of Saül, French Guiana, with the goal of producing a manual of the vascular plant flora, has yielded a new species ofSmilax. A diagnostic characteristic that differentiates this new species from all other species ofSmilax known from the Guianas and the Amazonian region is the admedial ramified branching pattern of the tertiary leaf veins.